The present invention concerns novel approaches for preparation by synthesis of the 3-phosphate derivatives of 1D-1-(1',2'-di-O-fattyacyl-sn-glycero-3'-phospho)-myo-inositols (PtdIns), referred to as the D-3-phosphorylated phosphoinositides or the 3-PPI (FIG. 1), their structural and stereochemical analogues, and, key starting materials and intermediates of these approaches.
3-PPI are relatively new members of the phosphoinositide group of cellular lipids with emerging critical roles in intracellular signalling. Synthetic 3-PPI and analogues are needed as reagents for defining their biological functions, and for developing diagnostics and therapeutics.
The 3-PPI (FIG. 1) including phosphatidylinositol-3-phosphate, PtdIns(3)P, and the bis- and tris-phosphate derivatives PtdIns(3,4)P.sub.2 and PtdIns(3,4,5)P.sub.3, have been found in eukaryotic cells (1), and the occurrence of PtdIns(3,5)P.sub.2 has been suggested (2). These compounds have been demonstrated as activators of protein kinase C isoforms .delta., .epsilon., and .sup.n (3), and are putative messengers in cellular signal cascades pertinent to inflammation, cell proliferation, transformation, protein kinesis, and cytoskeletal assembly (4). Minute quantities are found in cells and biochemical studies to determine the cellular targets of the 3-PPI, their metabolic fate, and their roles in the cell cycle have been handicapped because 3-PPI have not been available. Methods for synthesis of 3-PPI have been sought recently (5). These prior art methods suffer from some unique and common problems related respectively to the choice of starting materials for the myo-inositol as well as the diacylglycero-lipid moieties in the 3-PPI. In contrast with the present invention, all start with sn-1,2-diacylglycerols as the lipid moiety in the 3-PPI, and consequently are prone to problems of poor chemical stability endemic to 1,2-diacylglycerols. The latter isomerize readily via neighboring O-acyl migration to equilibrium mixtures comprising the 1,2-, 1,3- and 2,3-diacylglycerols (6). This equilibration is tantamount to racemization which is virtually complete for sn-1,2-di(short-chain)fattyacylglycerols. Therefore, resulting 3-PPI may contain racemic 1,2-2,3- and 1,3-difattyacyl structures, especially with hexanoyl and related short-chain fattyacyls.